Proliferation and differentiation of cells within multicellular organisms is controlled by hormones and polypeptide growth factors. These diffusable molecules allow cells to communicate with each other and act in concert to form organs, and to repair and regenerate damaged tissue. Examples of hormones and growth factors include the steroid hormones (e.g., estrogen, testosterone), parathyroid hormone (PTH), follicle stimulating hormone (FSH), the interleukins, platelet derived growth factor (PDGF), epidermal growth factor (EGF), granulocyte-macrophage colony stimulating factor (GM-CSF), erythropoietin (EPO), insulin and calcitonin.
Hormones and growth factors influence cellular metabolism by binding to receptors. Receptors may be integral membrane proteins that are linked to signalling pathways within the cell, such as second messenger systems. Other classes of receptors are soluble molecules, such as certain transcription factors.
Insulin belongs to a group of protein/polypeptide hormones. Insulin increases the rate of synthesis of glycogen, fatty acids, and proteins and stimulates glycolysis and cell proliferation. It also promotes the transport of glucose, and some other sugars, and amino acids into muscle and fat cells. The mature form of insulin consists of a 30 amino acid residue B chain, that is at the N-terminus of the propeptide form, and a 21 amino acid residue A chain, that is C-terminal. Proinsulin also contains a connecting peptide between the B chain and A chain that is cleaved out to form mature insulin. The B chain and A chain are covalently joined by two disulfide bonds. The B-chain, C-peptide, A-chain motif is found in several other proteins, including relaxin (U.S. Pat. No. 4,835,251), insulin-like growth factors (IGF) I and II (Bang and Hall, In "Insulin-like Growth Factors", P. N. Schofield (eds.), 151-177, Oxford University Press, Oxford, 1992), Leydig Factor (Bullesbach et al., J. Biol. Chem. 270:16011-16015, 1995, and early placenta insulin-like factor (EPIL; Chassin et al., Genomics 29:465-470, 1995). Unlike other members of the insulin superfamily, IGF I and IGF II have D and E domains that are cleaved post-translationally. Cysteines that are involved in disulfide bonds are conserved in all the memebers of the family and play a role in the tertiary structure of the molecules.
Another member of the insulin-family, relaxin, was recognized from crude extracts of sow corpora lutea (Hisaw, Proc. Soc. Exp. Biol. Med. 23:661, 1962 and Fevold et al., J. Am. Chem. Soc. 52:3340, 1930) and known to be involved in pregnancy and parturition. Relaxin has been identified in humans, and found to most abundant in the corpora lutea of pregnancy, but is also found in male seminal fluid (Weisse, Ann. Rev. Physiolog. 46:43-52, 1984). The relaxins of various species are divergent, with only 50% or less sequence homology between porcine, rat, shark and human relaxins. Within human species, there are two relaxin isoforms that have been identified (Hudson et al., Nature 301:628, 1983 and U. S. Pat. No. 4,758,516), however, only one of the mRNA from the isoforms (H2) has been identified in corpora lutea.
Like proinsulin, the B chain-C peptide-A chain motif is recapitulated in the relaxins. Preprorelaxin and preproinsulin both have a signal sequence, followed by the B chain, C peptide, A chain. The mature molecule of both proteins has the signal peptide and C peptide removed with the B and A chains joined by both inter- and intra-chain disulfide bonds (James et al., Nature 267:544-546, 1977). It has been postulated that the relaxins play roles in ripening of the cervix, thickening of the endometrium and increasing vascularization of the pregnant uterus and enhancing motility of sperm (U.S. Pat. No. 4,835,251). In addition, relaxin H2 was been found to bind in rat heart and has been tested for treatment of heart failure (U.S. Pat. No. 5,478,807).
Potential therapeutics and tools for identification of therapeutics that exhibit some tissue specificity are very valuable. Members of the insulin family are known to be potent growth factors. Therefore, a novel member of this family, with some specificity would be very valuable for understanding and regulating diseases associated with expression of these proteins.